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Isosilybin B causes androgen receptor degradation in human prostate carcinoma cells via PI3K-Akt-Mdm2-mediated pathway

Abstract

The identification and development of novel nontoxic phytochemicals that target androgen and androgen receptor (AR) signaling remains a priority for prostate cancer (PCA) control. In the present study, we assessed the antiandrogenic efficacy of isosilybin B employing human PCA LNCaP (mutated AR), 22Rv1 (mutated AR) and LAPC4 (wild-type AR) cells. Isosilybin B (10–90 μM) treatment decreased the AR and prostate specific antigen (PSA) levels in LNCaP, 22Rv1 and LAPC4 cells, but not in non-neoplastic human prostate epithelial PWR-1E cells. Isosilybin B treatment also inhibited synthetic androgen R1881-induced nuclear localization of AR, PSA expression and cell growth, and caused G1 arrest. In mechanistic studies identifying AR degradation, isosilybin B caused increased phosphorylation of Akt (Ser-473 and Thr-308) and Mdm2 (Ser-166), which was linked with AR degradation as pretreatment with PI3K inhibitor (LY294002)-restored AR level. Further, overexpression of kinase-dead Akt largely reversed isosilybin B mediated-AR degradation suggesting a critical role of Akt in AR degradation. Antibody pull-down results also indicated that isosilybin B treatment enhances the formation of complex between Akt, Mdm2 and AR, which promotes phosphorylation-dependent AR ubiquitination and its degradation by proteasome. Together, present findings identify a novel mechanism for isosilybin B-mediated anticancer effects in human PCA cells.

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References

  • Bernard D, Pourtier-Manzanedo A, Gil J, Beach DH . (2003). Myc confers androgen-independent prostate cancer cell growth. J Clin Invest 112: 1724–1731.

    Article  CAS  Google Scholar 

  • Chen CD, Welsbie DS, Tran C, Baek SH, Chen R, Vessella R et al. (2004). Molecular determinants of resistance to antiandrogen therapy. Nat Med 10: 33–39.

    Article  Google Scholar 

  • Chen L, Gilkes DM, Pan Y, Lane WS, Chen J . (2005). ATM and Chk2-dependent phosphorylation of MDMX contribute to p53 activation after DNA damage. EMBO J 24: 3411–3422.

    Article  CAS  Google Scholar 

  • Chen S, Xu Y, Yuan X, Bubley GJ, Balk SP . (2006). Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1. Proc Natl Acad Sci USA 103: 15969–15974.

    Article  CAS  Google Scholar 

  • Cinar B, De Benedetti A, Freeman MR . (2005). Post-transcriptional regulation of the androgen receptor by mammalian target of rapamycin. Cancer Res 65: 2547–2553.

    Article  CAS  Google Scholar 

  • Davis-Searles PR, Nakanishi Y, Kim NC, Graf TN, Oberlies NH, Wani MC et al. (2005). Milk thistle and prostate cancer: differential effects of pure flavonolignans from Silybum marianum on antiproliferative end points in human prostate carcinoma cells. Cancer Res 65: 4448–4457.

    Article  CAS  Google Scholar 

  • Deep G, Oberlies NH, Kroll DJ, Agarwal R . (2007). Isosilybin B and isosilybin A inhibit growth, induce G1 arrest and cause apoptosis in human prostate cancer LNCaP and 22Rv1 cells. Carcinogenesis 28: 1533–1542.

    Article  CAS  Google Scholar 

  • Deep G, Singh RP, Agarwal C, Kroll DJ, Agarwal R . (2006). Silymarin and silibinin cause G1 and G2-M cell cycle arrest via distinct circuitries in human prostate cancer PC3 cells: a comparison of flavanone silibinin with flavanolignan mixture silymarin. Oncogene 25: 1053–1069.

    Article  CAS  Google Scholar 

  • Dhanalakshmi S, Agarwal C, Singh RP, Agarwal R . (2005). Silibinin up-regulates DNA-protein kinase-dependent p53 activation to enhance UVB-induced apoptosis in mouse epithelial JB6 cells. J Biol Chem 280: 20375–20383.

    Article  CAS  Google Scholar 

  • Gioeli D, Black BE, Gordon V, Spencer A, Kesler CT, Eblen ST et al. (2006). Stress kinase signaling regulates androgen receptor phosphorylation, transcription, and localization. Mol Endocrinol 20: 503–515.

    Article  CAS  Google Scholar 

  • Grossmann ME, Huang H, Tindall DJ . (2001). Androgen receptor signaling in androgen-refractory prostate cancer. J Natl Cancer Inst 93: 1687–1697.

    Article  CAS  Google Scholar 

  • Haag P, Bektic J, Bartsch G, Klocker H, Eder IE . (2005). Androgen receptor down regulation by small interference RNA induces cell growth inhibition in androgen sensitive as well as in androgen independent prostate cancer cells. J Steroid Biochem Mol Biol 96: 251–258.

    Article  CAS  Google Scholar 

  • Hershko A, Ciechanover A . (1998). The ubiquitin system. Annu Rev Biochem 67: 425–479.

    Article  CAS  Google Scholar 

  • Jemal A, Siegel R, Ward E, Murray T, Xu J, Thun MJ . (2007). Cancer statistics, 2007. CA Cancer J Clin 57: 43–66.

    Article  Google Scholar 

  • Kim NC, Graf TN, Sparacino CM, Wani MC, Wall ME . (2003). Complete isolation and characterization of silybins and isosilybins from milk thistle (Silybum marianum). Org Biomol Chem 1: 1684–1689.

    Article  CAS  Google Scholar 

  • Klein KA, Reiter RE, Redula J, Moradi H, Zhu XL, Brothman AR et al. (1997). Progression of metastatic human prostate cancer to androgen independence in immunodeficient SCID mice. Nat Med 3: 402–408.

    Article  CAS  Google Scholar 

  • Lee DK, Chang C . (2003a). Endocrine mechanisms of disease: expression and degradation of androgen receptor: mechanism and clinical implication. J Clin Endocrinol Metab 88: 4043–4054.

    Article  CAS  Google Scholar 

  • Lee HJ, Chang C . (2003b). Recent advances in androgen receptor action. Cell Mol Life Sci 60: 1613–1622.

    Article  CAS  Google Scholar 

  • Liao Z, Wang S, Boileau TW, Erdman Jr JW, Clinton SK . (2005). Increased phospho-AKT is associated with loss of the androgen receptor during the progression of N-methyl-N-nitrosourea-induced prostate carcinogenesis in rats. Prostate 64: 186–199.

    Article  CAS  Google Scholar 

  • Lin HK, Wang L, Hu YC, Altuwaijri S, Chang C . (2002). Phosphorylation-dependent ubiquitylation and degradation of androgen receptor by Akt require Mdm2 E3 ligase. EMBO J 21: 4037–4048.

    Article  CAS  Google Scholar 

  • Lin HK, Yeh S, Kang HY, Chang C . (2001). Akt suppresses androgen-induced apoptosis by phosphorylating and inhibiting androgen receptor. Proc Natl Acad Sci USA 98: 7200–7205.

    Article  CAS  Google Scholar 

  • Mendoza N, Phillips GL, Silva J, Schwall R, Wickramasinghe D . (2002). Inhibition of ligand-mediated HER2 activation in androgen-independent prostate cancer. Cancer Res 62: 5485–5488.

    CAS  PubMed  Google Scholar 

  • Miyamoto H, Altuwaijri S, Cai Y, Messing EM, Chang C . (2005). Inhibition of the Akt, cyclooxygenase-2, and matrix metalloproteinase-9 pathways in combination with androgen deprivation therapy: potential therapeutic approaches for prostate cancer. Mol Carcinog 44: 1–10.

    Article  CAS  Google Scholar 

  • Muller MM, Schreiber E, Schaffner W, Matthias P . (1989). Rapid test for in vivo stability and DNA binding of mutated octamer binding proteins with ‘mini-extracts’ prepared from transfected cells. Nucleic Acids Res 17: 6420.

    Article  CAS  Google Scholar 

  • Nagabhushan M, Miller CM, Pretlow TP, Giaconia JM, Edgehouse NL, Schwartz S et al. (1996). CWR22: the first human prostate cancer xenograft with strongly androgen-dependent and relapsed strains both in vivo and in soft agar. Cancer Res 56: 3042–3046.

    CAS  PubMed  Google Scholar 

  • Nelson WG, De Marzo AM, Isaacs WB . (2003). Prostate cancer. N Engl J Med 349: 366–381.

    Article  CAS  Google Scholar 

  • Paajarvi G, Roudier E, Crisby M, Hogberg J, Stenius U . (2005). HMG-CoA reductase inhibitors, statins, induce phosphorylation of Mdm2 and attenuate the p53 response to DNA damage. FASEB J 19: 476–478.

    Article  Google Scholar 

  • Pan Y, Chen J . (2003). MDM2 promotes ubiquitination and degradation of MDMX. Mol Cell Biol 23: 5113–5121.

    Article  CAS  Google Scholar 

  • Pliskova M, Vondracek J, Kren V, Gazak R, Sedmera P, Walterova D et al. (2005). Effects of silymarin flavonolignans and synthetic silybin derivatives on estrogen and aryl hydrocarbon receptor activation. Toxicology 215: 80–89.

    Article  CAS  Google Scholar 

  • Reddy GP, Barrack ER, Dou QP, Menon M, Pelley R, Sarkar FH et al. (2006). Regulatory processes affecting androgen receptor expression, stability, and function: potential targets to treat hormone-refractory prostate cancer. J Cell Biochem 98: 1408–1423.

    Article  CAS  Google Scholar 

  • Seidlova-Wuttke D, Becker T, Christoffel V, Jarry H, Wuttke W . (2003). Silymarin is a selective estrogen receptor beta (ERbeta) agonist and has estrogenic effects in the metaphysis of the femur but no or antiestrogenic effects in the uterus of ovariectomized (ovx) rats. J Steroid Biochem Mol Biol 86: 179–188.

    Article  CAS  Google Scholar 

  • Sharifi N, Farrar WL . (2006). Androgen receptor as a therapeutic target for androgen independent prostate cancer. Am J Ther 13: 166–170.

    Article  Google Scholar 

  • Singh RP, Agarwal R . (2002). Flavonoid antioxidant silymarin and skin cancer. Antioxid Redox Signal 4: 655–663.

    Article  CAS  Google Scholar 

  • Singh RP, Agarwal R . (2006). Prostate cancer chemoprevention by silibinin: bench to bedside. Mol Carcinog 45: 436–442.

    Article  CAS  Google Scholar 

  • Tong Q, Zhao J, Chen Z, Zeng F, Lu G . (2003). Effects of blocking androgen receptor expression with specific hammerhead ribozyme on in vitro growth of prostate cancer cell line. Chin Med J (Engl) 116: 1515–1518.

    CAS  Google Scholar 

  • Wang L, Lin HK, Hu YC, Xie S, Yang L, Chang C . (2004). Suppression of androgen receptor-mediated transactivation and cell growth by the glycogen synthase kinase 3 beta in prostate cells. J Biol Chem 279: 32444–32452.

    Article  CAS  Google Scholar 

  • Wellington K, Jarvis B . (2001). Silymarin: a review of its clinical properties in the management of hepatic disorders. BioDrugs 15: 465–489.

    Article  CAS  Google Scholar 

  • Wen Y, Hu MC, Makino K, Spohn B, Bartholomeusz G, Yan DH et al. (2000). HER-2/neu promotes androgen-independent survival and growth of prostate cancer cells through the Akt pathway. Cancer Res 60: 6841–6845.

    CAS  PubMed  Google Scholar 

  • Yang L, Xie S, Jamaluddin MS, Altuwaijri S, Ni J, Kim E et al. (2005). Induction of androgen receptor expression by phosphatidylinositol 3-kinase/Akt downstream substrate, FOXO3a, and their roles in apoptosis of LNCaP prostate cancer cells. J Biol Chem 280: 33558–33565.

    Article  CAS  Google Scholar 

  • Zhou BP, Liao Y, Xia W, Zou Y, Spohn B, Hung MC . (2001). HER-2/neu induces p53 ubiquitination via Akt-mediated MDM2 phosphorylation. Nat Cell Biol 3: 973–982.

    Article  CAS  Google Scholar 

  • Zi X, Agarwal R . (1999). Silibinin decreases prostate-specific antigen with cell growth inhibition via G1 arrest, leading to differentiation of prostate carcinoma cells: implications for prostate cancer intervention. Proc Natl Acad Sci USA 96: 7490–7495.

    Article  CAS  Google Scholar 

  • Zi X, Feyes DK, Agarwal R . (1998). Anticarcinogenic effect of a flavonoid antioxidant, silymarin, in human breast cancer cells MDA-MB 468: induction of G1 arrest through an increase in Cip1/p21 concomitant with a decrease in kinase activity of cyclin-dependent kinases and associated cyclins. Clin Cancer Res 4: 1055–1064.

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Tyler N Graf, MS for preparative isolation of the pure flavonolignans used in these studies. This work was supported by NCI RO1 CA104286 (to DJK and RA).

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Correspondence to R Agarwal.

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Deep, G., Oberlies, N., Kroll, D. et al. Isosilybin B causes androgen receptor degradation in human prostate carcinoma cells via PI3K-Akt-Mdm2-mediated pathway. Oncogene 27, 3986–3998 (2008). https://doi.org/10.1038/onc.2008.45

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